CN1668293A - Methods of preventing or delaying the manifestation of cancer using artemisinin-like compounds - Google Patents

Abstract

The present invention provides methods for preventing or delaying the progression of cancer by administering a free radical generator to a subject. Representative free radical generators include endoperoxide compounds such as endoperoxide-containing sesquiterpene compounds such as artemisinin and its analogs, ativudine and its analogs, 1, 2, 4-trioxanes and 1, 2, 4, 5-tetraoxanes. Intracellular iron concentrations can be increased by administration of iron salts or complexes.

Description

Methods of preventing or delaying the manifestation of cancer using artemisinin-like compounds

Cross References to Related Applications

This application claims the benefit of U.S. Provisional Application No. 60/386,928, filed June 6, 2002, under 35 U.S.C. 119.

Field of Invention

The present invention relates to methods of preventing or delaying the manifestation of cancer by administering a free radical generating agent to a subject.

Background of the Invention

Artemisinins are sesquiterpene lactones isolated from the plant Artemisia annua L., an extract of which has been used to treat malaria for at least 1600 years. Artemisinin molecules contain endoperoxide bridges, which react with iron atoms to form free radicals. The antimalarial effect of artemisinin is due to its reaction with the heme in the parasite to form free radicals, resulting in cell death. Cancer cells have significantly higher iron influx than normal cells. Thus, artemisinin and artemisinin analogs have been shown to be cytotoxic to established tumors and tumor cell lines (see, for example, Woerdenbag et al. (1993) J. Nat. Prod. 56(6) : 849-56; Lai & Singh (1995) "Cancer Lett." (Cancer Lett.) 91: 41-6; Efferth et al. (2001) "International Oncology Journal" (Int.J.Oncol.) 18: 767-73; Li et al. (2001) "Bioorganic Medicinal Chemistry Communication" (Bioorg.Med.Chem.Lett.) 11: 5-8; Singh&Lai (2001) "Life Science" (Life Sci.) 70: 49-56; Efferth et al. ( 2002) "Biochem.Pharmacol." (Biochem.Pharmacol.) 64: 617-23; Efferth et al. (2002) "Blood Cells, Molecules & Diseases" (Blood Cells, Molecules & Diseases) 28(2): 160-8; Sadava et al. (2002) Cancer Lett. 179:151-6).

Many analogs of artemisinin and other biologically active compounds containing endoperoxide bridges have been described (see, for example, U.S. Patent No. 5,180,840; U.S. Patent No. 5,216,175; U.S. Patent No. 5,225,427; (1998) "Journal of Medicinal Chemistry" (J Med.Chem.) 41 (6): 952-64; Posner et al. (1999) "Journal of Medicinal Chemistry" (J Med.Chem.) 42: 300-4; Li et al. ( 2001) "Bioorganic Medicinal Chemistry Letters" (Bioorg.Med.Chem.Lett.) 11: 5-8; Wu et al. (2001) "European Journal of Medicinal Chemistry" (Eur.J.Med.Chem.) 36: 469- 79; Posner et al. (2003) J Med. Chem. 46:1060-5). Artemisinin analogs that have been used to treat malaria include dihydroartemisinin, artemether, artesunate, artether, dihydroartemisinin propyl carbonate, and artelinic acid.

Artemisinin is a relatively safe drug with few and minimal side effects even at high doses. Oral doses of 70 mg/kg/day for 6 days have been used to treat malaria in humans. After treating cancer patients with artesunate, no significant adverse side effects were observed (oral dose 50 mg/day; intramuscular dose 60 mg/day; for 9 months) (Singh & Verma (2002) "Oncology Journal" (Arch . Oncol.) 10(4):279-80). Artemisinin and artemisinin analogs have also been used to treat skin disorders such as psoriasis, blistering dermatoses, viral warts, mulluscum conntagiosum, and hemorrhoids (see, for example, U.S. Patent No. 4,978,676 ; US Patent No. 5,219,880). Artemisinin and artemisinin analogs have also been used to prevent malaria.

Cancer generally arises from the tendency of single cells to proliferate in an uncontrollable manner. There is a need in the art for methods of inhibiting cells prone to develop cancer in order to prevent or delay the onset of cancer. The present invention fulfills this need.

Summary of Invention

The present invention provides methods for preventing or delaying the manifestation of cancer. The method includes administering to the subject an amount of a free radical generator effective to prevent or delay the manifestation of cancer. In certain embodiments, the free radical generator is a compound containing an endoperoxygen bridge. Generally, the endoperoxide compound is selected from the group consisting of sesquiterpene lactones and their alcohols, carbonates, esters, ethers and sulfonates, atyfrine, 1,2,4 - the group consisting of trioxanes and 1,2,4,5-tetraoxanes. Described endoperoxide compound (endoperoxide compound) can be the compound of following general formula or its pharmaceutically acceptable salt:

R₁为氢、羟基、烷基或具有下列通式：where R is or

_R is hydrogen, hydroxyl, alkyl or has the following general formula:

或

-OR ₂ ,

or

wherein R ₂ is alkyl or aryl and n is 1-6.

Representative endoperoxide compounds useful in the practice of the present invention include, but are not limited to, artemisinin, dihydroartemisinin, artemether, artether, artesunate, atriptyline acid, and dihydroartemisinin Propyl carbonate.

Free radical generators also include compounds that do not contain an endoperoxo bridge, but can react with iron to form free radicals, such as carbon-based free radicals. The source of the free radical generator can be natural (eg, isolated from a plant), synthetic or semi-synthetic. For example, free radical generators can be produced by expressing enzymes in the relevant synthetic pathways in microbial hosts (see, e.g., Martin et al. (2003) Nature Biotechnol. Published online: June 1, 2003 , doi: 10.1038/nbt833).

In certain embodiments, the method further comprises the step of administering to the subject an effective amount of an intracellular iron-enhancing agent. Typical intracellular iron-fortifying compounds are iron salts and complexes, including iron bile salts, ferroglycine sulfate, dextran iron complex, iron peptone, iron sorbitol, iron oxide, With sugary iron, whole ferritin and holotransferrin.

The methods of the invention are applicable to any mammalian subject, such as a human subject. Thus, in certain embodiments, carcinogen-exposed subjects treated with a free radical generating agent according to the methods of the invention remain cancer-free twice as long as control animals not treated with the free radical generating agent.

In addition, the methods of the invention reduce the likelihood of cancer manifesting. Thus, in certain embodiments, none of the carcinogen-exposed subjects treated with a free radical generating agent according to the methods of the present invention developed cancer, compared to 43% of control animals not treated with a free radical generating agent. cancer.

In another aspect, the invention provides a kit comprising a free radical-generating agent and instructions for using the free radical generating agent to delay or prevent the manifestation of cancer in a subject. In certain embodiments, the kits of the invention may further include an iron fortifier.

Brief description of attached drawings

The above aspects of the present invention and many additional advantages are more apparent when considered in conjunction with the accompanying drawings, as also can be better understood with reference to the following detailed description, in which:

Figure 1 shows tumor development from exposure to carcinogens as described in Example 1, but without treatment for 1-38 weeks (indicated as "◆") or after weekly oral administration of artemisinin (indicated as "■") Graphical representation of percentage of animals.

Accompanying drawing 2 shows after being exposed to carcinogens as described in Example 2, but after 1-10 weeks without treatment (indicated as "◆") or after treatment with artemisinin mixed in food (indicated as "■") ) Graphical representation of the percentage of animals developing tumors.

A detailed description of the preferred implementation

In one aspect, the present invention provides a method of preventing or delaying the development of cancer, the method comprising administering to a subject an amount of a free radical generator effective to prevent or delay the development of cancer. The methods of the invention are applicable to any mammalian subject, such as a human subject or an animal subject. The term "preventing or delaying the development of cancer" as used herein refers to preventing or delaying the development of cancer that would occur without the administration of the free radical generating agent of the present invention. The term "cancer manifestation" refers to the point in time when a cancer becomes clinically manifest or detectable by any method suitable for the detection and diagnosis of cancer. Thus, cancer manifestation can be delayed for a defined period of time or delayed indefinitely. For example, the manifestation of cancer can be prevented or delayed by killing precancerous cells or cells that are genetically or environmentally predisposed to abnormal or uncontrolled cell proliferation, by preventing these cells from dividing, or by preventing the abnormal or uncontrolled division of these cells. By preventing or delaying the manifestation of cancer, free radical generating agents can prolong the time before and/or reduce the likelihood of cancer manifesting.

The term "free radical generator" refers to any agent that can react with iron or iron-containing compounds to generate free radicals, such as carbon-based free radicals. In certain embodiments, the free radical generator is an endoperoxide compound. The term "endoperoxide compound" refers to a compound containing an endoperoxy bridge which reacts in the presence of iron and iron-containing compounds to form free radicals. Endoperoxide compounds can also form free radicals in the presence of copper and manganese. Representative endoperoxide compounds are described above, however, it will be apparent that other endoperoxide compounds may also be used for this purpose.

Typical endoperoxide compounds useful in the practice of this invention are artemisinin and its analogs, as well as other compounds containing endoperoxide bridges that can react with iron and iron-containing compounds to form free radicals. Thus, suitable endoperoxide compounds include sesquiterpene lactones and their alcohols, carbonates, esters, ethers and sulfonates, atyfrine, 1,2,4-trioxane Classes and 1,2,4,5-tetraoxanes. Suitable sesquiterpene compounds of the present invention containing endoperoxides include compounds of the following formula and pharmaceutically acceptable salts thereof:

或

R₁为氢、羟基、烷基或具有下列通式：where R is

or

_R is hydrogen, hydroxyl, alkyl or has the following general formula:

或

-OR ₂ ,

or

Wherein R ₂ is an alkyl group or an aryl group, and n is 1-6. The term "alkyl" as used herein refers to a lower alkyl group containing 1-6 carbon atoms, preferably 1-4 carbon atoms. The alkyl groups of the present invention may be straight chain or branched chain groups. The term "aryl" refers to monocyclic and polycyclic aromatic groups containing 4-14 backbone carbon or heteroatoms, including carbocyclic aryl and heterocyclic aryl groups. A carbocyclic aryl group is an aryl group in which all ring atoms are carbon. A heterocyclic aryl group contains 1 to 4 heteroatoms as ring atoms, the remaining ring atoms being carbon. Representative aryl groups include, for example, phenyl and benzyl. Pharmaceutically acceptable salts include alkali metal or alkaline earth metal salts, preferably sodium or potassium salts.

二氢青蒿素(R₁＝-OH)；artesunicacid(R₁＝-OCO(CH₂)₂CO₂H)；以及青蒿琥酯；蒿甲醚(R₁＝-OCH₃)；和蒿乙醚(R₁＝-OC₂H₅)。本发明其它有代表性的内过氧化物化合物包括阿替林酸、二氢青蒿素丙基碳酸酯、阿替夫林(Ro.42-1611)及其类似物(Biirgen等(1994)Sixth Int.Cong.Infect.Dis.Abst.427，p.152，Prague)、1，2，4-三噁烷类(Peters等(1993)Ann.Trop.Med.Parasit.87(1)：9-16)和1，2，4，5-四噁烷类(Vennerstrom等(1992)《药物化学杂志》(J Med Chem.)35(16)：3023-3027)。青蒿素的其它合适的结构类似物描述在例如美国专利US5,216,175和US5,180,840、Cumming等(1998)《药物化学杂志》(J Med Chem.)41(6)：952-64和PCT专利申请WO97/01548、WO 99/33461和WO 00/42046中。For example, endoperoxide compounds of the present invention include: artemisinin, wherein R is

Dihydroartemisinin (R ₁ =-OH); artesunicacid (R ₁ =-OCO(CH ₂ ) ₂ CO ₂ H); and artesunate; artemether (R ₁ =-OCH ₃ ); Diethyl ether (R ₁ =-OC ₂ H ₅ ). Other representative endoperoxide compounds of the present invention include atriptyline acid, dihydroartemisinin propyl carbonate, atiefrine (Ro. 42-1611) and analogs thereof (Biirgen et al. (1994) Sixth Int.Cong.Infect.Dis.Abst.427, p.152, Prague), 1,2,4-trioxanes (Peters et al. (1993) Ann.Trop.Med.Parasit.87 (1): 9- 16) and 1,2,4,5-tetraoxanes (Vennerstrom et al. (1992) J Med Chem. 35(16):3023-3027). Other suitable structural analogs of artemisinin are described, for example, in U.S. Patent Nos. 5,216,175 and 5,180,840, Cumming et al. Applications WO 97/01548, WO 99/33461 and WO 00/42046.

The amount of free radical generator effective in inhibiting cells susceptible to developing cancer is up to the maximum tolerated dose, but the concentration is not critical and can vary widely. The precise amount will of course vary with the compound, the route of administration, the physical condition of the patient and other factors. The daily dose can be administered as a single dose or it can be divided into multiple doses.

The amount of free radical generator actually administered may be a prophylactically effective amount, which term is used herein to mean the amount required to substantially prevent or delay the development of cancer. Effective doses may be extrapolated from dose-response curves derived from in vitro or animal model test systems. Animal models are generally used to determine ideal dosage ranges and routes of administration. Such information can be used to determine useful doses and routes of administration for human or other mammalian administration. Determination of effective doses is well within the purview of those skilled in the art. Therefore, the actual dosage administered depends on the specific subject, and it is preferable to optimize the dosage so as to obtain the desired effect without significant side effects.

The prophylactic efficacy and possible toxicity of free radical generators, such as endoperoxide compounds, can be determined in cell culture or experimental animals by standard pharmaceutical procedures (e.g. _ED50 , the dose effective to treat 50% of the population; and _LD50 , Even the dose that kills 50% of the population). The dose ratio between therapeutic and toxic effects is the therapeutic index and it can be expressed as the ratio between _LD50 and _ED50 . Free radical generators that exhibit high therapeutic indices, eg, a therapeutic index of at least about 60, are particularly suitable for practicing the methods of the invention. The data obtained from cell culture assays and animal studies can be used in determining a range of dosage for use in humans or other mammals. The dosage of free radical generators lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage will generally vary within this range, depending upon the dosage form employed, the sensitivity of the subject and the route of administration. Therefore, the optimal amount may vary depending on the method of administration, and will generally be that of a commonly used agent administered in the same or similar form. For example, for topical or oral administration, generally 1-3 times per day can be performed.

For endoperoxide compounds such as artemisinin and its analogs, about 0.1 to about 20 mg/kg body weight/day, such as about 0.5 to about 15 mg/kg body weight/day, for example about 1 to about 10 mg Good results were obtained with formulations comprising the compound at a level per kg body weight per day. Typical daily oral doses of endoperoxides, such as artemisinin, artemether or artether, are from about 20 mg to about 200 mg, such as from about 40 mg to about 180 mg or from about 80 mg to about 160 mg, for adult subjects . Similar dosages can be used for other free radical generators.

The free radical generators of the present invention may be administered as described in Examples 1 and 2 by any effective route, for example by oral administration. Free radical generators can also be administered parenterally. Methods of administration include topical, inhalation, buccal, intraarterial, subcutaneous, intramedullary, intravenous, intranasal, intrarectal, intraocular and other commonly used methods. The free radical generator can be formulated into a composition which additionally contains a suitable pharmaceutically acceptable carrier, including excipients and other compounds which facilitate administration of the free radical generator to a mammalian subject . More specific techniques for formulation and administration can be found in the latest edition of "Remington's Pharmaceutical Sciences" (Maack Publishing Co, Easton PA).

Free radical generators may be formulated for oral administration as described in Examples 1 and 2 using pharmaceutically acceptable carriers well known in the art, in dosages suitable for oral administration. Such carriers enable the free radical generator to be formulated as tablets, pills, lozenges, capsules, liquids, gels, syrups, slurries, suspensions and the like, suitable for ingestion by a subject. The free radical generator for oral use can be formulated with, for example, a solid excipient, the resulting mixture optionally ground, and the mixture of granules processed, after adding suitable other compounds, if desired, to prepare tablets or lozenges Drug core. Suitable excipients include carbohydrate or protein fillers. They include, but are not limited to: sugars, including lactose, sucrose, mannitol, or sorbitol; starches from corn, wheat, rice, potatoes, or other plants; celluloses, such as methylcellulose, hydroxypropylmethyl cellulose or sodium carboxymethylcellulose; and gums, such as acacia and tragacanth; and proteins, such as gelatin and collagen. If desired, a disintegrating or solubilizing agent may be added, such as cross-linked polyvinylpyrrolidone, agar, alginic acid or a salt thereof, such as sodium alginate.

The tablet cores are coated with suitable coatings, such as concentrated sugar solutions, which may also contain gum arabic, talc, polyvinylpyrrolidone, carbomer gel, polyethylene glycol and/or titanium dioxide, lacquer solutions and suitable organic coatings. solvent or solvent mixture. Dyestuffs or pigments may be added to the tablets or dragee coatings for product identification or to characterize the amount of active compound (ie, dosage).

For example, free radical generators for oral administration can be formulated as push-fit capsules made of gelatin, as well as soft, sealed capsules consisting of gelatin and a coating such as glycerol or sorbitol. The push-fit capsules can contain the endoperoxide compound in admixture with a filler or binders, such as lactose or starches; lubricants, such as talc or magnesium stearate; and, optionally, stabilizers. In soft capsules, the free radical generators may be dissolved or suspended in suitable liquids, with or without stabilizers, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. For example, a free radical generator, such as an endoperoxide compound, such as artemether, artether, artemisinin or other artemisinin analogues can be dissolved in an oil, such as soybean oil, olive oil or peanut oil. Suitable soybean oils include, but are not limited to, soybean oil from Glycine Soya Bentham (eg Shanghai Number 2 Oil Factory, cat. no. 91102).

Compositions for parenteral administration include aqueous solutions of one or more free radical generators. For injection, the endoperoxide compounds of the invention may be formulated in aqueous solutions, such as physiologically compatible buffers, such as Hank's solution, Ringer's solution, or physiologically buffered saline. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Additionally, suspensions of free radical generators may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include: fatty oils, such as sesame oil; or synthetic fatty acid esters, such as ethyl oleate or triglycerides; or liposomes. The suspension may also optionally contain suitable stabilizers or agents which increase the solubility of the free radical generators to allow for the preparation of highly concentrated solutions.

For topical or nasal administration, penetrants suitable to penetrate the particular barrier will generally be used in the formulation. Examples thereof are 2-pyrrolidone, N-methyl-2-pyrrolidone, dimethylacetamide, dimethylformamide, propylene glycol, methanol or isopropanol, dimethylsulfoxide and laurocapram. Other agents may also be included to render the formulation cosmetically acceptable. Examples thereof are fats, waxes, oils, dyes, fragrances, preservatives, stabilizers and surfactants. Keratolytic agents such as those known in the art may also be included. Examples are salicylic acid and sulfur. For topical administration, the composition may be in the form of a transdermal ointment or patch for systemic delivery of the compound, and may be prepared in conventional manner (see, e.g., Barry, Dermatological Formulations (Drugs and the Pharmaceutical Sciences--Dekker); Harrys Cosmeticology (Leonard Hill Books).

For rectal administration, the compositions may be administered in the form of suppositories or retention enemas. Such compositions can be prepared by mixing the free radical generator with a suitable non-irritating excipient which is solid at ordinary temperatures but liquid at the rectal temperature, whereby the Melts in the rectum to release the drug. Suitable excipients include, but are not limited to, cocoa butter and polyethylene glycols.

The respective amounts of these different types of additives will be apparent to those skilled in the art, the optimal amounts being the same as in other known formulations designed for the same type of administration. For example, for stratum corneum penetration enhancers, concentrations of about 0.1% to about 15% are generally included.

Compositions containing free radical generators of the present invention can be prepared in a manner similar to that known in the art (for example, by conventional mixing, dissolving, granulating, tableting, milling, emulsifying, encapsulating, entrapping or lyophilizing). Law). The composition may also be modified by conventional means (eg, coatings) to provide suitable release characteristics, eg sustained or targeted release.

Compositions containing free radical generators can be prepared in the form of salts and can be salted with many acids including, but not limited to, hydrochloric acid, sulfuric acid, acetic acid, lactic acid, tartaric acid, malic acid, succinic acid, and the like. Salts are more soluble in aqueous or other protic solvents than the corresponding free base forms.

After such compositions formulated to contain the free radical generator and an acceptable carrier have been prepared, they can be placed in a suitable container and labeled for use.

The free radical generators of the invention may be administered alone or in combination with one or more other therapeutically active agents. For example, higher efficacy of endoperoxide compounds can be achieved by increasing oxygen tension, reducing antioxidant uptake, and blocking peroxidase and catalase with drugs such as miconazole. The effectiveness of free radical generators can also be enhanced by administering intracellular iron-fortifying compounds. Accordingly, certain embodiments of the present invention also include intracellular iron fortifiers. Free radical generators, such as endoperoxide compounds, react with iron to form free radicals. It has been shown that administration of iron salts or the iron-carrying protein holotransferrin increases the sensitivity of cancer cells to artemisinin or dihydroartemisinin (Lai & Singh (1995) Cancer Lett. 91:41 -46; Moore et al. (1995) Cancer Lett. 98:83-7; Singh & Lai (2001) Life Sci. 70:49-56; Sadava et al. (2002) Cancer Lett. 1179:151-6). According to the methods of some embodiments of the present invention, administration of an intracellular iron fortifier increases the effectiveness of the free radical generating agent in delaying or preventing the manifestation of cancer.

The term "intracellular iron booster" refers to any active agent effective to increase the intracellular concentration of ferrous iron in the body, including pharmaceutically acceptable iron salts and iron complexes. Iron salts useful in the practice of the present invention include ferrous fumarate, ferrous sulfate, ferrous carbonate, ferrous citrate, ferrous gluconate, and ferrous lactate. Iron complexes useful in the practice of the present invention generally include: pharmaceutically acceptable complexes containing iron, such as iron bile salts, ferrous sulfate glycine, iron dextran complexes, iron peptone, iron sorbitol, iron oxide and sugar-containing iron; and iron complexed with iron-binding proteins and glycoproteins, such as holoferritin and holotransferrin.

Concentrations of intracellular iron fortifiers used in the present invention will generally be up to the maximum tolerated dose for the particular subject and agent, which will vary with the agent, subject, disease condition, and other factors, as described above. vary with each other. A dosage range of about 1 to about 20 mg iron/kg subject body weight/day is generally used for this purpose. In one embodiment, the intracellular iron fortifier may be administered prior to administration of the free radical generator.

The methods of the invention provide a significant delay in the manifestation of cancer in a subject. Thus, in certain embodiments, as described in Example 1, a carcinogen-exposed subject treated with a free radical generating agent according to the methods of the present invention remains cancer-free for at least as long as without treatment with the free radical generating agent. approximately 2 times that of control animals.

Furthermore, the methods of the invention reduce the likelihood of cancer manifesting. Thus, in certain embodiments, as described in Examples 1 and 2, subjects treated with free radical generators exposed to carcinogens had fewer carcinogens than control animals not treated with endoperoxide compounds. Cancer will occur. In certain embodiments, as described in Example 2, exposure to carcinogens treated with a free radical generating agent according to the methods of the invention compared to 43% of cancers in control animals not treated with a free radical generating agent None of the subjects developed cancer.

In another aspect, the invention provides kits comprising one or more free radical generators and instructions for using the free radical generators to delay or prevent the manifestation of cancer. The instructions may indicate conditions under which administration of a free radical generator may be particularly beneficial, such as family history of cancer, pre-existing cancer, or exposure to carcinogens. The instructions also provide advice on administration, such as dosage. In certain embodiments, the kits of the present invention may further include instructions for iron fortification agents and their use in combination with endoperoxide compounds.

The following examples merely illustrate the best mode currently known for carrying out the invention and should not be used to limit the invention.

Example 1

This example describes a representative method for preventing or delaying the development of cancer in carcinogen-exposed rats by weekly oral intubation administration of artemisinin.

Materials and methods: Female Sprague-Dawley rats were administered a single intragastric dose of the carcinogen, 7,12-dimethylbenzo[ a] Anthracene (DMBA; Sigma Chemicals, St Louis, MO; 50 mg/kg body weight). Animals were randomized into two treatment groups, one group receiving weekly oral intubation of artemisinin (Holley Pharmaceuticals, Fullerton, CA; 10 mg/kg in 10 mg/ml of olive oil) or olive oil (control group). Artemisinin treatment was initiated one week after DMBA administration. Mammary tumors present in animals were palpated weekly. The time and location of detection and size are recorded for each tumor present in the animal.

Results: Six weeks after carcinogen treatment, none of the artemisinin-treated animals developed tumors, while 17% of the control animals had tumors. Ten weeks after carcinogen treatment, tumors developed in 9% of artemisinin-treated animals compared with 58% of control animals. Twenty weeks after carcinogen treatment, 54% of artemisinin-treated animals developed tumors, compared with 83% of control animals. Thirty-eight weeks after carcinogen treatment, 64% of artemisinin-treated animals developed tumors, compared with 100% of control animals. These results are shown in Table 1 and Figure 1 of the accompanying drawings.

Table 1. Tumor development in animals treated weekly with artemisinin week % of animals with tumors present   Artemisinin treatment group control group 6 0 17 7 9 25 9 9 42 10 9 58 11 9 67 12 27 67 13 45 67 16 45 75 17 45 83 20 54 83 29 64 92 31 64 100 38 64 100

Thus, weekly treatment with artemisinin delayed tumor expression in carcinogen-treated animals. In addition, fewer animals developed tumors than the control group.

Example 2

This example describes a representative method for preventing or delaying the onset of cancer in carcinogen-exposed rats by providing artemisinin in admixture with food.

Materials and Methods: Female rats were similarly treated with the carcinogen DMBA as described in Example 1 . Rats were randomly divided into two treatment groups. Artemisinin (0.05%) was mixed into the diet of one group of animals the day after DMBA treatment. The daily intake of artemisinin was estimated to be approximately 10 mg/kg/day based on daily food intake. Another group of animals (control group) was given chow periodically.

Results: After 10 weeks of carcinogen treatment, none of the artemisinin-fed animals developed tumors, while 43% of the animals in the control group had tumors, as shown in Table 2 and accompanying drawing 2.

Table 2. Tumor occurrence in animals fed artemisinin week % of animals with tumors present The group fed artemisinin control group 6 0 9 7 0 13 8 0 26 9 0 39 10 0 43

While preferred embodiments of the present invention have been illustrated and described, it will be understood that various changes may be made therein without departing from the spirit and scope of the invention.

Claims (19)

1. A method for preventing or delaying the appearance of cancer, comprising administering to a subject a free radical generator in an amount effective for preventing or delaying the appearance of cancer. 2. The method of claim 1, wherein said free radical generator is an endoperoxide compound selected from the group consisting of sesquiterpene lactones and alcohols, carbonates, esters, ethers thereof And sulfonates, atifrin, 1,2,4-trioxanes and 1,2,4,5-tetraoxanes. 3. The method of claim 2, wherein said endoperoxide compound is a compound of the following general formula and pharmaceutically acceptable salts thereof:

where R is

or Wherein _R is hydrogen, hydroxyl, alkyl or has the following general formula: -OR ₂ , or

Wherein R ₂ is an alkyl group or an aryl group, and n is 1-6. 4. The method of claim 2, wherein said endoperoxide compound is a sesquiterpene compound selected from the group consisting of artemisinin, dihydroartemisinin, artemether, artether, artesunate, Group consisting of atriptyline acid and dihydroartemisinin propyl carbonate. 5. The method of claim 4, wherein said endoperoxide compound is artemisinin. 6. The method of claim 4, wherein the endoperoxide compound is artemether. 7. The method of claim 2, wherein about 0.1 to about 20 mg/kg of the endoperoxide compound is administered to the subject per day. 8. The method of claim 2, wherein the endoperoxide compound is administered orally or topically. 9. The method of claim 1, wherein the free radical generator is artemisinin, the subject is a human subject, and artemisinin is orally administered in powder, tablet or capsule form, The dose is about 0.1-20 mg/kg/day. 10. The method of claim 1, wherein administering the free radical generating agent to the subject provides a delay in the development of the cancer. 11. The method of claim 1, wherein administering the free radical generating agent to the subject reduces the likelihood of the cancer developing. 12. The method of claim 1, further comprising administering to the subject an effective amount of an intracellular iron fortifier. 13. The method of claim 12, wherein the intracellular iron fortifier is selected from the group consisting of iron bile salt, ferrous sulfate glycine, iron dextran complex, iron peptone, iron sorbitol, iron oxide, sugar-containing The group consisting of iron, holoferritin, and holotransferrin. 14. The method of claim 12, wherein the intracellular iron fortifier is administered to the subject prior to administration of the free radical generating agent. 15. A kit comprising a free radical generator and instructions for using the free radical generator to prevent or delay the manifestation of cancer. 16. The kit of claim 15, wherein the free radical generator is an endoperoxide compound selected from the group consisting of sesquiterpene lactones and alcohols, carbonates, esters, ethers and A group consisting of sulfonates, atifrin, 1,2,4-trioxanes and 1,2,4,5-tetraoxanes. 17. The kit of claim 16, wherein said endoperoxide compound is artemisinin. 18. The kit of claim 16, wherein the endoperoxide compound is artemether. 19. The kit of claim 15, further comprising an iron fortifier and instructions for using the iron fortifier.

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